Amino acid derivatives shown by the below formula (I)
(wherein R1 means a hydrogen atom or a straight chain or branched chain alkyl groups with 1-5 carbon atoms. Hereinafter it is same.) have a wide variety of use as a base or a base substance for introducing a functional molecule in case of synthesizing a monomer unit for synthesis of Boc-type PNA, an amino acid derivative introducing a Boc-type functional molecule and the like.
In particular, as shown in FIG. 1, PNA (Peptide Nucleic Acid) has the structure in which the sugar phosphoric acid skeleton in a natural nucleic acid such as DNA is converted into the N-(2-aminoethyl)glycine skeleton, is high in a double strand formation performance and a base sequence recognition performance compared with a natural nucleic acid, further is intact for an in vivo nuclease and protease, and therefore its application to a gene therapy as an antisence molecule is examined, attracting attention in recent years. The above characteristics of PNA is due to the fact that the sugar phosphoric acid skeleton in a natural nucleic acid has negative charge in neutral conditions whereby an electrostatic repulsion between complementary chains is produced, and in contrast in PNA having N-(2-aminoethyl) glycine skeleton without charge no electrostatic repulsion is produced between complementary chains.
Synthesis of PNA is carried out by sequentially combining an amino acid (especially glycine) derivative (monomer unit) which is introduced by any one of four kinds of bases (A, T(U), C and G) constituting DNA or RNA according to an aimed base sequence using a conventional solid peptide synthesis method. As for monomer units for the synthesis of PNA, there are two types, Fmoc type and Boc type, as shown in FIG. 2 (B represents base), though synthetic methods of monomer units are established and use of a Fmoc type which, enables to utilize a general DNA automatic synthesis machine to synthesize a PNA oligomer currently has become a major trend. However, because in case of synthesizing PNA by use of a Boc type monomer unit there is an advantage that a functional molecule unstable in basic conditions can be introduced in PNA, establishment of a PNA synthesis method using a Boc type monomer unit becomes an urgent matter.
As one of obstacles to hinder establishment of a PNA synthesis method using a Boc type monomer unit, it can be cited that a simple and cheap synthesis method for a monomer unit before introduction of a base, that is, a Boc type amino acid derivative shown in the formula (I), is not to be established. Also, because an amino acid derivative of the formula (I) has use as a base substance for introducing other functional molecule in stead of the base, synthesis of an amino acid derivative introducing a functional molecule becomes easy if its simple and cheap synthesis method is established.
The synthetic method for an amino acid derivative of the formula (I) usually makes ethylenediamine a starting material and contains a step to introduce t-butoxycarbonyl group (Boc) to one nitrogen atom and a step to introduce —CHR1—COOH to the other nitrogen.
As a method to obtain t-butoxycarbonylaminoethylamine by introduction of Boc to one nitrogen atom of ethylenediamine, for example, (1) methods are reported in which t-butoxycarboxylic acid anhydride is directly reacted to ethylenediamine in a reaction solvent such as chloroform, methanol or dioxane (J. Med. Chem., 38(22), 4433-8; 1995, Bull. Korean Chem. Soc., 15(12), 1025-7; 1994, Eur. J. Med. Chem., 26(9), 915-20; 1991, Synth. Commun., 20(16), 2559-64; 1990, Aust. J. Chem., 39(3), 447-55; 1986),

and (2) a method in which t-butoxycarboxylic acid anhydride is converted to an active ester followed by reaction with ethylenediamine (JP, A 11-012234,).

Also, as a method to obtain an amino acid derivative of the formula (I) by introduction of —CHR1—COOH to t-butoxycarbonylaminoethylamine, a method is reported in which benzyl group is introduced to the unprotected nitrogen atom of t-butoxycarbonylaminoethylamine, (3) followed by reaction with benzyl bromoacetate and then by the catalytic reduction (J. Org. Chem., 62(2), 411-416; 1997).

Further, as a method to obtain an amino acid derivative of the formula (I) by introduction of Boc to the other nitrogen atom of the ethylenediamine derivative in which —CHR1—COOH is introduced to one nitrogen atom, (4) a method is reported in which t-butoxycarboxylic acid anhydride is reacted to N-(2-aminoethyl)glycine (Heimer, E. P.; Gallo-Torres, H. E.; Felix, A. M.; Ahmad, M.; Lambros, T. J.; Scheidl, F.; Meienhofer, J. Int. J. Pept. Protein Res. 23(2), 203-211, 1984).

However, as a method to prepare t-butoxycarbonylaminoethylamine, in the method 1 the aimed substance can be obtained in relatively good yield, though di(t-butoxycarbonylamino) ethylene and t-butoxycarboxylic acid are produced as byproducts, existing in a reaction solvent such as chloroform, methanol or dioxane. Owing to this, a partition extraction procedure or partition chromatography are necessary, making it difficult to prepare t-butoxycarbonylaminoethylamine efficiently in a large amount and cheaply.
Also, although the method 2 has an advantage that di(t-butoxycarbonylamino)ethylene is not produced as a byproduct, the total yield is as low as about 60% due to a multistep reaction, and because used reagents must be removed by partition chromatography, it is difficult to prepare t-butoxycarbonylaminoethylamine efficiently in a large amount and cheaply just like the method 1.
Therefore, both methods 1 and 2 are inappropriate as a method to industrially prepare t-butoxycarbonylaminoethylamine.
Also, as a method to obtain an amino acid derivative of the formula (I) from t-butoxycarbonylaminoethylamine, the method 3 is a multistep reaction, and a partition extraction procedure is necessary, being inappropriate for an industrial preparation.
Further, as a method to obtain an amino acid derivative of the formula (I), the method A has an advantage that partition chromatography is unnecessary, though the yield is around 60%, being inappropriate for an industrial preparation. That is, an efficient method to obtain a photo-functional PNA molecule has not been established due to a low efficiency in the synthesis of an amino acid derivative of the formula (I). Therefore, needed are a method to obtain an amino acid derivative of the formula (I) and the development of the amino acid derivative to make a more efficient synthesis of the photo-functional PNA molecule possible in case of using an amino acid derivative of the formula (I).